Alagawany (2021) Early life thermal stress modulates hepatic expression of thermotolerance related genes and physiological responses in two rabbit breeds,

Early life heat stress negatively affects rabbit production and well‐being. However, the physiological response to acute heat stress in later life is not clearly defined. The present study aims to investigate the effects of early and late heat stress at 36°C on some blood constituents, antioxidant enzymes activity in the blood, and muscle in New Zealand white and Baladi Black rabbits. A total of sixty post‐weaning rabbits of each breed were randomly divided into two groups; control groups (NZWC and BBC) and early heat‐stressed groups for six hours at 36 ± 1°C and 62% relative humidity (RH) (NZWT and BBT groups). After heat stress, six rabbits from each group were slaughtered for blood and muscle tissue collection. The surviving rabbits were kept at 28 ± 1°C and 40% RH till 13 weeks of age. At the end of 13 weeks, all rabbits were exposed to late heat stress as precious described to perform four groups: single late stressed groups; NZWC2, BBC2, and double stressed groups; NZWT2 and BBT2. After late heat stress, six rabbits from each group were slaughtered for blood and muscle tissue collection. The early and late heat stress caused a significant reduction in the blood creatine kinase, lactate dehydrogenase, and high‐density lipoprotein and antioxidant enzymes' activity in blood and muscle of both NZW and BB rabbits compared with the control groups. While, the blood total cholesterol, triglycerides, total lipids levels, and lipid peroxidation activity in blood and muscle were significantly increased due to the early and late heat‐stressed both breeds compared with the control groups. It could be concluded that the early heat stress at 36°C has negative effects on several physiological indicators and antioxidant activities in the blood and muscle of NZW and BB rabbits.

Background: Sperm-mediated gene transfer (SMGT) is a technique that utilizes the ability of the spermatozoa to take up exogenous DNA. Growth hormone is anabolic hormone that plays an important role in muscle-building process and milk production in all animals. High blood concentration of growth hormones (GH) was observed for animals that were genetically selected for high milk production or for low carcass fatness levels. The present study aimed to investigate and enhance the capacity of ovine spermatozoa to uptake exogenous growth hormone cDNA and its impact on sperm motility. The current study is an introduction for further future studies to produce transgenic Egyptian sheep characterized with high productive performance. Methods: The growth hormone cDNA sequence was extracted from pituitary gland of Egyptian × Holstein (EH_ GH) cattle and subcloned into the pmKate2-N vector to construct the EH_GH-pmKate2-N expression vector. The complete sequence of EH_GH mRNA was registered in GenBank (AC: KP221576). A total of three groups were assessed for the sperm uptake experiment, namely, negative control, positive control, and dimethyl sulfoxide (DMSO) groups; all treated groups were incubated with the EH_GH-pmKate2-N vector. The expression of EH_GH protein was detected in DH10B cells using a fluorescence microscope and the SDS polyacrylamide gel electrophoresis. Results: The EH_GH-pmKate2-N vector was expressed in cultured Escherichia coli cells, and the molecular weight of EH_GH protein was 24,558 Da. The EH_GH-pmKate2-N vector was introduced efficiently into the heads of the spermatozoa in the DMSO and positive control groups. Incubation of the spermatozoa with the vector caused a significant reduction in progressive motility compared to the negative control. Conclusion: The present results demonstrated the ability of ovine spermatozoa to take up the exogenous vector without notable deleterious effects on sperm motility. In subsequent studies, the successful introduction of the exogenous GH expression vector into the sperm head allows for the production of GH-transgenic sheep characterized by a high growth rate in order to reduce the meat shortage in Egypt.

This study aims to produce transgenic ovine spermatozoa bearing Ossimi sheep growth hormone (Os_GH) cDNA using different methods. The complete coding sequence of Os_GH has been registered in GenBank accession no. KP221575. The sequence of Os_GH cDNA has been subcloned into pmkate2-N expression vectors to construct Os_GH-pmKate2-N vector. Five groups of sperm uptake were submitted. All groups were incubated at 37 °C for 1 h: Control (sperm cells were incubated without vector), Traditional incubation (sperm cells were incubated with vector), Heat shock (sperm cells were incubated with vector at 4 °C for 20 min and heated for 2 min at 42 °C), Heat shock + Dimethyl sulfoxide (DMSO) (sperm cells were incubated with vector and supplemented with 3% of DMSO and then submitted to heat shock regime) and DMSO (sperm cells were incubated with vector and supplemented with 3% DMSO). The sperm genomic DNA in groups was extracted. The Os_GH-pmKate2-N vector was introduced efficiently into the head of sperm cells in all treated groups. Adding DMSO either with or without heat shock increased the sperm uptake. The progressive motility was reduced (P < 0.05) by 29.9% in heat shock group compared to the control. Adding DMSO improved (P < 0.05) the total and progressive motilities by 8.2% and 19.8%, respectively in heat shock group compared to the heat shock group without DMSO. The results documented the ability of ovine spermatozoa to uptake the exogenous vector. Also, sperm incubation with 3% DMSO is the best method to introduce the exogenous vector into spermatozoa without notable adverse effects on sperm motilities.

Background: Identification of molecular characterization of genes underlying livestock productive traits may allow applying advanced biotechnology techniques to improve animal productivity. Growth hormone (GH) controls body growth rate, milk production, reproduction as well as carbohydrate, lipid, and protein metabolism. Therefore, the present study aims to investigate the genetic variations of growth hormone cDNA sequences between Assaf sheep (As_GH) and Boer goat (Bo_GH) that mainly used for genetic improvement in Egypt using bioinformatics analysis. Growth hormone cDNA was isolated from the pituitary gland tissue of Assaf sheep Boer goat and subcloned into pTZ57R/T cloning vector for sequencing. Results: Molecular weight of As_GH cDNA was 665 bp and was 774 bp for Bo_GH cDNA. The complete coding sequences (CDS) of As_GH and Bo_GH were registered in the GenBank database under accession number (AC: MH128986 and AC: MG744290, respectively). High homology percentage was observed (99.5%) between AS_GH and Bo_GH protein sequences with one different amino acid in the As_GH protein sequence (Arg 194). The protein sequence of As_GH has only one motif signature; Somatotropin_1 from 79 to 112 aa compared to Bo_GH protein sequences and GenBank database that had two motifs signature. The growth hormone cDNA sequence of Assaf sheep has a unique three single nucleotide polymorphisms (SNPs) (A 637 A 638 G 639) that encodes for arginine (Arg 194); this insertion mutation (AAG) was not found in the growth hormone cDNA sequences of Boer goat in the present study and GenBank database breeds. This mutation can be used to develop SNPs markers for Assaf sheep. Conclusions: GH sequence of Assaf and Boer goat is highly conserved and the homogeny in the codon region (99.5%). The Assaf sheep GH sequence has a unique three SNPs that may be used to develop SNPs markers for such breed. Further studies are needed to investigate the genetic variations of growth hormone gene in different sheep and goat breeds in Egypt and document the relationship between these variations and the productive performance of animals.